Reactive Oxygen Species Are Key Mediators of Demyelination in Canine Distemper Leukoencephalitis but not in Theiler’s Murine Encephalomyelitis

(1) Background: Canine distemper virus (CDV)-induced demyelinating leukoencephalitis (CDV-DL) in dogs and Theiler’s murine encephalomyelitis (TME) virus (TMEV)-induced demyelinating leukomyelitis (TMEV-DL) are virus-induced demyelinating conditions mimicking Multiple Sclerosis (MS). Reactive oxygen species (ROS) can induce the degradation of lipids and nucleic acids to characteristic metabolites such as oxidized lipids, malondialdehyde, and 8-hydroxyguanosine. The hypothesis of this study is that ROS are key eector molecules in the pathogenesis of myelin membrane breakdown in CDV-DL and TMEV-DL. (2) Methods: ROS metabolites and antioxidative enzymes were assessed using immunofluorescence in cerebellar lesions of naturally CDV-infected dogs and spinal cord tissue of TMEV-infected mice. The transcription of selected genes involved in ROS generation and detoxification was analyzed using gene-expression microarrays in CDV-DL and TMEV-DL. (3) Results: Immunofluorescence revealed increased amounts of oxidized lipids, malondialdehyde, and 8-hydroxyguanosine in CDV-DL while TMEV-infected mice did not reveal marked changes. In contrast, microarray-analysis showed an upregulated gene expression associated with ROS generation in both diseases. (4) Conclusion: In summary, the present study demonstrates a similar upregulation of gene-expression of ROS generation in CDV-DL and TMEV-DL. However, immunofluorescence revealed increased accumulation of ROS metabolites exclusively in CDV-DL. These results suggest dierences in the pathogenesis of demyelination in these two animal models

Hypertensive nonobstructive hydrocephalus as main magnetic resonance imaging feature in a dog with disseminated choroid plexus carcinomatosis

Obstructive or nonobstructive hypertensive hydrocephalus is reported in choroid plexus tumors. Choroid plexus tumors typically present as T2‐weighted hyperintense intraventricular masses with occasional cerebrospinal fluid‐drop metastasis. Acquired neoplastic nonobstructive hydrocephalus without visible mass lesion in magnetic resonance imaging is not reported in dogs. A 4.5‐year‐old Rhodesian Ridgeback presented with reduced mental status, unilaterally absent pupillary light reflex, and neck pain. Magnetic resonance imaging revealed a nonobstructive hydrocephalus and widened lumbar subarachnoid space with no evidence of a primary mass lesion. Postmortem examination confirmed a disseminated choroid plexus tumor affecting the ependyma and choroid plexi of all ventricles and the cerebral and lumbar subarachnoid space. Disseminated choroid plexus carcinomatosis should be considered as a possible cause of hypertensive hydrocephalus even in absence of a primary mass

RALGAPA1 Deletion in Belgian Shepherd Dogs with Cerebellar Ataxia.

Several genetically distinct forms of cerebellar ataxia exist in Belgian shepherd dogs. We investigated a litter in which two puppies developed cerebellar ataxia. The clinical signs stabilized at around six weeks of age, but remained visible into adulthood. Combined linkage and homozygosity mapping delineated a 5.5 Mb critical interval. The comparison of whole-genome sequence data of one affected dog to 929 control genomes revealed a private homozygous ~4.8 kb deletion in the critical interval, Chr8:14,468,376_14,473,136del4761. The deletion comprises exon 35 of the RALGAPA1 gene, XM_038544497.1:c.6080-2893_6944+1003del. It is predicted to introduce a premature stop codon into the transcript, truncating ~23% of the wild-type open reading frame of the encoded Ral GTPase-activating protein catalytic subunit α 1, XP_038400425.1:(p.Val2027Glnfs*7). Genotypes at the deletion showed the expected co-segregation with the phenotype in the family. Genotyping additional ataxic Belgian shepherd dogs revealed three additional homozygous mutant dogs from a single litter, which had been euthanized at five weeks of age due to their severe clinical phenotype. Histopathology revealed cytoplasmic accumulation of granular material within cerebellar Purkinje cells. Genotyping a cohort of almost 900 Belgian shepherd dogs showed the expected genotype-phenotype association and a carrier frequency of 5% in the population. Human patients with loss-of-function variants in RALGAPA1 develop psychomotor disability and early-onset epilepsy. The available clinical and histopathological data, together with current knowledge about RALGAPA1 variants and their functional impact in other species, suggest the RALGAPA1 deletion is the likely causative defect for the observed phenotype in the affected dogs

Case report: Canine distemper virus infection as a cause of central nervous system disease in a Eurasian lynx (Lynx lynx)

The Eurasian lynx (Lynx lynx) represents an endangered species with only small populations remaining in Central Europe. Knowledge about the threat posed by potential infectious agents to these animals is crucial for informing ongoing protection measures. Canine distemper virus (CDV) is known to have a wide host range with infection reported in many mammalian species including several lynx species (Lynx pardinus, Lynx canadensis, Lynx rufus), but is an extremely rare finding in the Eurasian lynx. The present report describes a case of a Eurasian lynx showing central nervous signs, including apathy and ataxia. A CT scan revealed multiple hypodense areas in different localizations within the brain as well as enlarged liquid filled areas, leading to the suspicion of a degenerative process. Due to clinical deterioration, the animal was euthanized and submitted for macroscopical and histological investigations. Histological investigations revealed multifocal demyelinations in the cerebellum, brain stem and cervical spinal cord as well as a multifocal, perivascular, lymphohistiocytic meningoencephalitis. A CDV infection was confirmed by immunohistochemistry and RT-PCR analyses. This CDV infection of a Eurasian lynx resembles a classical chronic manifestation of distemper in dogs and highlights the threat posed by canine distemper to this species

Comparison of different in situ hybridization techniques for the detection of various RNA and DNA viruses

In situ hybridization (ISH) is a technique to determine potential correlations between viruses and lesions. The aim of the study was to compare ISH techniques for the detection of various viruses in different tissues. Tested RNA viruses include atypical porcine pestivirus (APPV) in the cerebellum of pigs, equine and bovine hepacivirus (EqHV, BovHepV) in the liver of horses and cattle, respectively, and Schmallenberg virus (SBV) in the cerebrum of goats. Examined DNA viruses comprise canine bocavirus 2 (CBoV-2) in the intestine of dogs, porcine bocavirus (PBoV) in the spinal cord of pigs and porcine circovirus 2 (PCV-2) in cerebrum, lymph node, and lung of pigs. ISH with self-designed digoxigenin-labelled RNA probe

An Equine Model for Vaccination against a Hepacivirus: Insights into Host Responses to E2 Recombinant Protein Vaccination and Subsequent Equine Hepacivirus Inoculation

Equine hepacivirus (EqHV) is the closest known genetic homologue of hepatitis C virus. An effective prophylactic vaccine is currently not available for either of these hepaciviruses. The equine as potential surrogate model for hepacivirus vaccine studies was investigated, while equine host responses following vaccination with EqHV E2 recombinant protein and subsequent EqHV inoculation were elucidated. Four ponies received prime and booster vaccinations (recombinant protein, adjuvant) four weeks apart (day −55 and −27). Two control ponies received adjuvant only. Ponies were inoculated with EqHV RNA-positive plasma on day 0. Blood samples and liver biopsies were collected over 26 weeks (day −70 to +112). Serum analyses included detection of EqHV RNA, isotypes of E2-specific immunoglobulin G (IgG), nonstructural protein 3-specific IgG, haematology, serum biochemistry, and metabolomics. Liver tissue analyses included EqHV RNA detection, RNA sequencing, histopathology, immunohistochemistry, and fluorescent in situ hybridization. Al-though vaccination did not result in complete protective immunity against experimental EqHV inoculation, the majority of vaccinated ponies cleared the serum EqHV RNA earlier than the control ponies. The majority of vaccinated ponies appeared to recover from the EqHV-associated liver insult earlier than the control ponies. The equine model shows promise as a surrogate model for future hepacivirus vaccine research

Influenza A (H10N7) virus causes respiratory tract disease in harbor seals and ferrets

Avian influenza viruses sporadically cross the species barrier to mammals, including humans, in which they may cause epidemic disease. Recently such an epidemic occurred due to the emergence of avian influenza virus of the subtype H10N7 (Seal/H10N7) in harbor seals (Phoca vitulina). This epidemic caused high mortality in seals along the north-west coast of Europe and represented a potential risk for human health. To characterize the spectrum of lesions and to identify the target cells and viral distribution, findings in 16 harbor seals spontaneously infected with Seal/H10N7 are described. The seals had respiratory tract inflammation extending from the nasal cavity to bronchi associated with intralesional virus antigen in respiratory epithelial cells. Virus infection was restricted to the respiratory tract. The fatal outcome of the viral infection in seals was most likely caused by secondary bacterial infections. To investigate the pathogenic potential of H10N7 infection for humans, we inoculated the seal virus intratracheally into six ferrets and performed pathological and virological analyses at 3 and 7 days post inoculation. These experimentally inoculated ferrets displayed mild clinical signs, virus excretion from the pharynx and respiratory tract inflammation extending from bronchi to alveoli that was associated with virus antigen expression exclusively in the respiratory epithelium. Virus was isolated only from the respiratory tract. In conclusion, Seal/H10N7 infection in naturally infected harbor seals and experimentally infected ferrets shows that respiratory epithelial cells are the permissive cells for viral replication. Fatal outcome in seals was caused by secondary bacterial pneumonia similar to that in fatal human cases during influenza pandemics. Productive infection of ferrets indicates that seal/H10N7 may possess a zoonotic potential. This outbreak of LPAI from wild birds to seals demonstrates the risk of such occasions for mammals and thus humans

The impact of upstream and downstream processing on the quality of oil bodies of partially de-hulled sunflower seeds

Few publications on oil bodies or oleosomes seem concerned about their quality (chemical and physical) ex-vivo. This work attempts to identify the main factors (processing and pre-processing) that affect the quality/integrity of sunflower seed oil bodies recovered through a wet-milling process. The physical state of seeds during wet milling had a significant impact on the quality of the oil body suspension. Pre-soaking for 6 hours before wet milling and multiple washing with alkaline buffer (0.1M sodium bicarbonate) was performed to isolate high quality oil body suspensions. It was evident from different physical measurements such as particle size, ζ-potential and light microscopy that pre-soaking had a positive influence on the quality of oil body suspensions with no significant signs of aggregation or coalescence. It was also observed that the resultant washed oil body suspensions were highly surface charged (-28.4 ± 1.2 mV) indicating very stable suspension phase behavior. Washing oil bodies not only removes non-integral, extraneous proteins (derived from the seed matrix) but enriches the lipid content including Tocopherol (α-tocopherol: 491.6 mg/kg of washed oil bodies compared with 252.6 mg/kg crude oil bodies). Changes in the composition of oil bodies after washing have been observed before, but this research also monitored the size of oil bodies after washing, and our results indicate that certain factors can shift the distribution of droplet size. It is believed that any change in average size of droplets indicate the presence of disrupted oil bodies whose surface chemistry has changed enough to compromise their integrity on washing. The retention of droplet size on washing may, therefore, be diagnostic for the recovery of intact oil bodies. An assessment of the integrity of oil bodies recovered from sunflower seeds after accelerated aging (5 months) was carried out. Free fatty acid was more pronounced in oil rather than oil bodies, this could be due to the elimination of some of the free acid bound to oil body during washing. Although some minor variation was observed during seed aging, however, the oil bodies remained stable in the final suspension. The results indicate that oil body membrane was extremely robust under extreme conditions and the integrity of oil bodies was preserved. In addition, oil bodies obtained in this study were resistant to oxidation due to the presence of naturally occurring antioxidants (including vitamin E) associated with them.. The results indicate that the physical barrier of surface membrane protein (oelosin) protect oil bodies against pro-oxidants